Single-cell transcriptomic profiling of cerebrospinal fluid reveals immune-cell heterogeneity and multicompartment MIF/CD74-related communication in tuberculous and purulent meningitis

Abstract

Background Tuberculous meningitis (TBM) is a severe central nervous system infection in which dysregulated host inflammation contributes to neurological injury. The cellular organization of the cerebrospinal fluid (CSF) immune microenvironment in TBM, particularly in comparison with other inflammatory meningitis conditions, remains incompletely defined. This study characterized CSF immune-cell heterogeneity in TBM and purulent meningitis (PM) using single-cell transcriptomic profiling. Methods Pre-treatment CSF immune cells from five patients with TBM and five patients with PM were profiled using 10x Genomics single-cell RNA sequencing. After quality control, ambient RNA assessment, and doublet filtering, 96,594 cells were retained. Major immune-cell populations and myeloid and granulocyte subclusters were annotated using canonical lineage markers and cluster-specific marker genes, with additional evaluation of doublet-related quality-control metrics and ambient RNA estimates. Immune-cell composition, CD4+ T-cell transcriptional states, macrophage migration inhibitory factor (MIF)/CD74 expression, CellChat-inferred communication, and in silico perturbation profiles were evaluated using donor-level comparisons where appropriate. Results Seven major CSF immune-cell populations were identified, including CD4+ T cells, CD8+ T cells, myeloid cells, granulocytes, B cells, plasma cells, and natural killer T (NKT)-like cells. Immune-cell composition varied markedly across donors. Several PM donors exhibited myeloid- or granulocyte-rich profiles, whereas several TBM donors showed greater CD4+ T-cell representation; group-level differences were exploratory and did not consistently reach statistical significance. Reannotation of the innate immune-cell compartment identified broad macrophage-like, monocyte-like, neutrophil-like, and CD1C+ dendritic cell–like populations. Two originally defined myeloid subclusters displayed coordinated T-cell-associated transcriptional programs and were interpreted as potentially misclassified or technically mixed cells rather than bona fide myeloid states. CD4+ T cells occupied a continuous transcriptional landscape comprising central memory (CM)-like, cytotoxicity/senescence-associated, and activated effector memory (EM)-like states with overlapping transcriptional programs. MIF was broadly detectable across immune-cell compartments, whereas CD74 was most prominent in antigen-presentation-associated populations. CellChat identified MIF-related communication in both disease groups, supporting a shared multicompartment inflammatory communication program rather than a TBM-specific pathway. Exploratory in silico perturbation prioritized MIF- and CD74-associated transcriptional programs for future validation. Conclusions This study provides a donor-aware single-cell transcriptomic atlas of the inflammatory CSF immune microenvironment in TBM and PM. The findings highlight substantial inter-individual heterogeneity, broad innate immune-cell patterns, overlapping CD4+ T-cell transcriptional states, and multicompartment MIF/CD74-related communication. These transcriptome-inferred findings are hypothesis-generating and require validation in larger cohorts using protein-level and functional approaches.